The third component of complement: covalent attachment of a radioactive sugar to the labile binding site of C3 via the alternative pathway.
A complement- (C) fixing particle consisting of agarose beads to which 5-thioglucose was attached by a --S--S-- bond (agarose-thioglucose) was employed to investigate the mechanism of attachment of C3 to surfaces. When whole serum containing [125I] C3 was incubated with agarose-thioglucose, labeled C3b was taken up in a form that was not removed by 2 M NaCl but was released by 10 mM dithiothreitol. Deposition of DTT-releasable C3b was dependent upon the alternative pathway of C activation. Gel electrophoresis of DTT-releasable C3b from similar experiments performed with unlabeled serum and agarose-[3H]thioglucose showed that the liberated C3b contained a molecule of radioactive thioglucose attached to the alpha'-chain by a covalent bond that was stable to mercaptoethanol. We propose that the thioglucose-alpha' chain bond was formed during the course of C activation by a reaction between the "labile binding site" of newly released C3b and the (then) particle-bound sugar. This formulation implies that the reaction by which C3b attaches to 5-thioglucose in this system is the reaction responsible for opsonization by C3b, and that the C3b-linked sugar represents a marker for the labile binding site. Incubation of the particle-bound C3b in serum resulted in the cleavage of the covalently linked alpha'-chain to several smaller polypeptides, the major cleavage product having a m.w. of 70,000.
D001665:Binding Sites; D003167:Complement Activation; D003176:Complement C3; D003179:Complement C3b; D003170:Complement Pathway, Alternative; D004229:Dithiothreitol / Q000494:pharmacology; D004492:Edetic Acid / Q000494:pharmacology; D004533:Egtazic Acid / Q000494:pharmacology; D005947:Glucose / Q000031:analogs & derivatives* / Q000378:metabolism; D006801:Humans; D008274:Magnesium / Q000378:metabolism; D012685:Sepharose / Q000378:metabolism
D003176:Complement C3; C003002:5-thio-D-glucose; D004533:Egtazic Acid; D003179:Complement C3b; D012685:Sepharose; D004492:Edetic Acid; D008274:Magnesium; D005947:Glucose; D004229:Dithiothreitol